During early fasting, increases in circulating pancreatic glucagon stimulate the gluconeogenic program in part via PKA-mediated phosphorylation and activation of the transcription factor CREB. The CREB coactivator, CRTC2, has been shown to function as an essential interacting partner in CREB-induced transcription activation. However, the molecular mechanisms by which CRTC2 selectively activates CREB target genes remain unclear. In an unbiased analysis of CREB- associated proteins, the protein arginine methyltransferase 5 (PRMT5) was identified as a putative interacting protein of CREB. Preliminary results indicate that overexpression of PRMT5 increases CREB activity and co-expression of CRTC2 further increases these effects, suggesting that PRMT5 functions as a coactivator. The proposed research is to determine whether PRMT5 modulates CREB-mediated gluconeogenic gene activation through arginine methylation of CREB or its coactivators in hepatic cells. In specific aim 1, the role of PRMT5 in methylating CREB and modulating the interaction between CREB and CRTC2 or CBP will be explored. CREB arginine residues that undergo methylation will be identified. In specific aim 2, the role of PRMT5 in mediating glucagon effects on the CREB-regulated gluconeogenic program will be determined by overexpression or depletion of PRMT5 in hepatocytes. The recruitment of PRMT5 to CREB-bound gluconeogenic promoters in response to glucagon will be revealed. The importance of arginine methylation in CREB-mediated transcription will be explored. In specific aim 3, the role of PRMT5 in regulating fasting glucose production will be evaluated. The potential effects of PRMT5 in promoting hyperglycemia will be tested under insulin resistance conditions. Collectively, the proposed studies will provide insight of how hepatic gluconeogenesis is regulated during fasting. Fasting hyperglycemia is common in insulin resistance patients in part due to up-regulation of hepatic gluconeogenesis. Thus, the results may lead to new molecular targets for the development of new therapeutic treatment of type II diabetes.